Apparatus for sample application in thin layer and paper chromatography

ABSTRACT

An apparatus for producing spots of specimens of material on a plate having a thin layer adsorbing surface, the specimens being dissolved in a diluting solvent for chromatographic analysis, with a plurality of units, each one of the units comprising a glass tube which is filled with a specimen through a funnel-like glass filling tube projecting perpendicularly from the upper surface of the specimen tube, the latter also having an open front and which is stoppered or otherwise covered by a teflon or other inert plug through which passes a capillary tube, the capillary tube having a bent front end protruding from the open front end of the glass tube so that the end of the capillary is very close to the adsorbing surface of the plate, and an air tube above the front end of the capillary which may be adjustably set to blow a low-pressure air stream onto the specimen emanating from the capillary to evaporate the solvent, thereby forming a spot of the specimen on the thin-layer adsorbing surface of the plate.

[451 Oct. 23, 1973 United States Patent [191 Rosenthal [54] APPARATUSFOR SAMPLE APPLICATION IN THIN LAYER AND PAPER CHROMATOGRAPHY wrs,mccmh.. ,rwth rmfnu ue SSVf n l0 .ego m.mSy .limmam C ewuum Wdmm fmdpTwnwwaa t Cwmmm Msmww nehv Tntls, S.Ia0.l BC uws Amsiw. O.mda ...VgnPa.ma rhec mbm wper. mam@ a .It wmmm nr h Amc k, r a l. P e i w mm H M,k w m m N wp n. Je .M d .h dy t nH m h kw R MUN r 09 u.Lh.r mY en n.heAN TC v .m A M M 7 7 Mmmm as ma 3. 76E-1884 SHEEY l UF 2 JMW/WMM@Jew/Paw `ATTORNEYS `one another on the adsorbent APPARATUS FOR SAMPLEAPPLICATION IN THIN LAYER AND PAPER CHROMATOGRAPHY BACKGROUND OF THEINVENTION The invention relates generally to medicomechanical apparatusand particularly to apparatus for simultaneously preparing a pluralityof specimens, especially for chromatographic analysis.

Various devices for producing spots for chromatographic analysis areknown, some being built on an experimental laboratory basis inuniversities, some others being commercially produced. The known devicesmay be divided into the two main groups, one to produce specimens instreak form and the other to form individual spots The objective inmaking streaks is usually purification of a mixture for preparativepurposes, while spot-making devices (as the presently described one) areused to facilitate qualitative or quantitative analysis of thechromatographically-separated materials. All prior art devices haveseveral features in common. Either they are hand-operated, which isslow, tedious work,.tying up laboratory personnel for the prep-V arationof the specimens and delaying the chromatographic analysis, or else theyemploy devices such as bellows, pumps, or other `motorized devices whichmake them bulky and/or expensive and may expose the sample solutions tocontamination from pump tubing. The non-motorized devices apply `liquidto the spots discretely rather than continuously; since the samples tobe analyzed are diluted by a solvent, it is obvious that the solvent ofeach one of these spots has to be evaporated before another portion ofsolution can be applied to the spot, until a comlete spot is formedwhich contains a sufficient quantity of material for the chormatographicanalysis. Another common feature found in the prior art is having eitherthe base-plate on which the spots are to be formed, or the pipet orcapillary, Amove to obtain more than one spot or streak. Variousoperations are known to fill the pipettes or capillaries, all of whichare cumbersome, V expensive and/or timeconsuming. All known devices havebeen built tobe operated by skilled personnel who have to devote theirVfull attention tothe process of preparing the spots, and sincethisoperation is strictly under manual control, it is more likelythannot that the sizes and thicknesses of the spots obtained, as well astheir positions relative to base, will not be uniform and consistent. l,Y i

The problems of timeand labor-saving production of the spots and of amore predictable and reliable conples are dissolved by a solvent (e.g.,chloroform) which has an adequate and desirable rate of evaporation.Specimens of the magnitude of l-2 ml are taken and deposited in the formof spots on the adsorbent surface by a continuous process, during whichthe solvent is continuously evaporated. The final spot contains,therefore, the entire quantity of the solids of the sample and yet doesnot have a size larger than 0.5 to 0.75 cm.

`The invention thus serves simultaneously to concentrate the specimenand apply it as a compact spot to a thin-layer plate, in preparation forchromatographic separation and analysis. Small, easily refillablereservoirs for the specimens are provided and capillary tubes are usedto take the solution out of the reservoirs for continuous deposit on thesupport plate on the same spot. A supply of air under low pressure,which may or may not be heated, is directed down by way of a nozzle ontop of the discharge end of the capillary tube. This flow oflow-pressure air envelops the end of the capillary without causing anair turbulence, evaporating at the same time the solvent and holding theforming spot to its desired diameter. This feature enables the arrangement of a plurality of capillaries side-by-side, which can thus beused simultaneously. Since the support plate with the adsorbent surfacelayer has the standard width of 8 inches, the arrangement of 8capillaries at distances of 1 inch from one another is easily feasible,without spots running into one another.

Objects and advantages of the invention will be set forth in parthereafter and in part will be obvious hereb from or may be learned bypracticing the invention, the

' consisting of ya sizable quantity of a sample junder sistency ofquality of these' spots, are intended to be i solved-by this invention.

SUMMARY oF THE nvvENTioN The invention consistsin such novel features,construction arrangements, combinations of parts and improvements as maybe shownand describedin connection with the apparatus herein disclosedby way of example only and as illustrative of a preferred embodi-`coninuous evaporation ofthe solvent of the sample.

A further object of the invention is to provide an apparatus by whichreliably reproducible results can be obtained without continuous manualoperation of the apparatus.

Furthermore, it is an object of rthe invention to provide an apparatuswhich can be operated expeditiously and without continuous supervision.

Yet another object `of the invention is to provide an apparatus which isof simple structure and can be eco nomically produced.

Various further and more specific purposes, features and advantages willclearly appear from the detailed description given below taken inconnection with the accompanying drawingwhich forms part of thisspecification and illustrates merely by way of example one embodiment ofthe apparatus of the invention.

BRIEF DESCRIPTION OF THE DRAWING In the following description and in theclaims, parts will be identified by specific names for convenience,

i but such names are intended to be as generic in their application tosimilar parts as the art will permit. Like reference characters denotelike parts in the figures of the drawing, in which FIG. 1 is anisometric perspective view of the apparatus of the invention;

FIG. 2 is a part of the apparatus having the reservoir tube for thespecimen and the support arm which the socket for the conventionalball-and-socket universal joint;

FIG. 3 shows the holder for the air tube, having the ball-shaped endconstituting the counterpart for the socket shown in FIG. 2.k

FIG. 4 shows the air tube with the flared nozzle end;

FIG. 5 is a plan view of the capillary tube which fits through theteflon plug which is seated firmly in the reservoir tube shown in FIG.2; and

FIG. 6 is a side view of the capillary shown in FIG. 5.

DESCRIPTION `OF THE PREFERRED EMBODIMENT Referring now in more detail tothe drawing illustrating a preferred embodiment by which the inventionmay be realized, there is the complete apparatus of the invention shownin FIGS. 1, identified by the general numeral 10.A The component partsare mounted on a base 11, preferably made of a corrosion-proof material.A plurality of vertical posts l2 (eight are shown in the Figure) arefixedly erected on base 1l, spaced apart in equal intervals inside-by-side relationship in a straight row. Conventional spring clips13 are attached to posts l2 to hold the support parts 14 for thetube-like reservoirs 15 in place. Support parts 14, which may be ofglass, comprise an upright member 16, the tube-like reservoir 15 affixedto the lower end of member 16 and inclined slightly'upward, anda supportarm y17 which is affixed to the upper end of member 16 at right anglethereto. The free end of support arm 17 is shaped as a socket 18 of auniversal ball joint,- into which the ballshaped e'nd 19`of the holder20 for the air tube 2l fits. Ball-shaped end 19 is the end of a shortarm 22 which is affixed to a tube 23 at right angle thereto." Ball end19 is held in place in socket 18 by a conventional spherical-jointspring clamp 24. Airtube 21 fits freely in glass tube 23 and is held inplace therein by short rubber sleeves'25 in` such manner that it iseasily adjust- 4 horizontal and has a short stopper 28 plugging thefront end thereof. The-stopper 28 consists preferably of a solid pieceof inert plastic (e.g. teflon or polyethylene) which securely coversthefront end of reservoir 15, but is removable. Capillary 31 passesthrough stopper 28 through an opening therein small enough to preventlateralmovement of capillary 3l in an air stream. This arrangement holdscapillary 31 securely in place. A small funnel-like tube 39 projectsfrom the upper surface of tube l5 just in front of its center throughwhich reservoir l5 is filled without taking it out of the apparatus.Capillary 31 consists preferably of a piece of stainless steelhypodermic needle material. A short piece of an inert plastic capillarytube 32 is attached to the front end of capillary 31. Plastic tube 32 isbent downward at right angle so that the end 33 thereof can be set veryclose to the adsorbent surface coating of the thin layer chromatographicplate 34 which is placed on base 1li underneath ends 33, without therisk of damaging this coating. Conventional thin-layer plate 34 consistsof a piece of thin glass or of a suitable plastic, the surface of whichis very evenly coated with an adsorbent material such as silica gel, inwhich the line of spots of the specimens to be chromatographicallyanaylzed is formed. Behind the line of posts l2 is a manifold 35 mountedon base ll, and air hoses 36 are attached to the upper ends of air tubes2l, leading therefrom to manifold 35 which is, in turn, connected by afeeding air hose 37 to a conventional source of compressed air under lowpressure. Laboratory-style Hoffman screw clamps 38 are arranged on airhoses 36 for a final adjustment of air supply to air tubes 2l. It is notabsolutely necessary to heat the air, but a provision may be made toincrease the temperature of the air above the ambient room temperatureby conventional heating means (e.g. by encirling the glass manifold witha heating coil, by directing infrared light at the line of formingspots, or by having the base 1l be the top surface of a hot plate).

The apparatus is operated as follows:

The support parts 14 are vertically adjusted on posts 12 so that ends 33of plastic capillaries 32 are very close to the coated surface of thechromatographic plate 34 which has been placed on base l1 so that therow of the ends 33 is approximately l inch away from the edge of theplate. Air tubes 2l are set vertically above the ends of the capillariesand are adjusted by the universal joint to a position where the flarednozzles 27 are above the ends of the capillaries. In the meantime,samples to be analyzed have been prepared by dissolving them in asolvent, for example, chloroform. Low pressure air is now turned on andthe flow of air is individually adjusted by means of hose clamps 38.Quantities of the diluted samples, generally 2 ml each, are filledthrough hole 39 into reservoir tubes l5. Since tubes 15 are slightlyinclined from the horizontal, the liquid will not flow onto the plasticStoppers 28, but will pass slowly by the capillary action throughcapillaries 31, and since ends 33 of the plastic capillaries 32 areslightly lower than the rear end of capillaries 3l in reservoir tubes15, a siphoning effect also takes place in feeding the liquid out of thereservoirs through the capillaries. The slow outflow of the samples willform relatively small spots as long as the air flowing out of flarednozzles will evaporate the, solvent instantaneously, continuouslypreventing thereby a spreading of the liquid in the absorbent coating.The air nozzles are eventually adjusted to a desired position so thatspots of approximately 0.5 to 0.75 cm diameter will result. The processcontinuesfully automatically until the entire contents of the reservoirsare spent and the solids of the samples are accumulated inthe spots.vThe reservoirs may be washed with fresh portions of solvent, if desired,and the wash solution added to the original spot by the same means usedin thel application of the latter. The plate with the spots thereon isnow taken off the base v and subjected to the conventionalchromatographic analysis. The accuracy of the automatedvprocess is fargreaterand more consistent than one of a tedious manual operation, andbetter comparable analytic results can be obtained.

It is understood that the present invention may also be used similarlyfor the application of sample spots for of the invention, that variousother changes and moditications may be made without departing from thespirit l of the invention.

What is claimed is:

l. A spot applicator unit for thin layer and paper chromatography, whichcomprises a. reservoir means for holding a measured amount of a liquidsample, said reservoir means havinginlet means for filling saidreservoir means and capillary tube outlet means for siphoning saidmeasured amount of liquid sample out of said reservoir means, saidcapillary tube outlet means having a discharge end outside the reservoirmeans and below the level of a liquid sample in said reservoir means;

b. air supply means for flowing a stream of low pressure air toevaporate solvent from said liquid sample siphoned out of said reservoirmeans; and

c. support means for supporting said reservoir means and said air supplymeans with said air supply means arranged to flow air over a liquidsample discharged from said discharge end, said support means includingadusting means for adjusting the position of said air supply meansrelative to said discharge end of said capillary outlet means thereby tocontrol the rate of evaporation of solvent from `said liquid samplesiphoned out of said reservoir means.

2. The applicator unit according to claim ll, wherein said air supplymeans has a flared outlet end positioned adjacent said discharge end,the position of said flared outlet end being adjustable by saidadjusting means.

3. `The applicator unit according to claim l2, whereinv said reservoirmeans has an elongated body portion, and said capillary outlet meanscomprises a capillary tube having a first part thereof inside saidelongated body portion and in communication with a liquid sample in saidelongated body portion, and said discharge end being provided by asecond part of the capillary tube positioned outside of saidvbodyportion and arranged transverse to said elongated body portion.

4. The applicator unit according` to claim 3, wherein f said second partof said capillary tube is substantially perpendicular'to said elongatedbody portion.

5. The applicator unit according to claim 3, wherein and said secondpart is of an insert plastic secured to said tirst part.

6. The applicator unit according to claim 3, wherein said capillary tubeis removably mounted in said elongated body portion.

7'. `The applicator unit according to claim 1, wherein said air supplymeans comprises air tiow controlv means.

, d. The applicator unit according to claim 1, wherein said air supplymeans comprises means for supplying heated air.

said tirst part of said capillary tube is of stainless steel having atirst part inside the reservoir meansand extending to the lowest part ofsaid reservoir means and a second part outside the reservoir means andextending below the level of said lowest part of said reservoir means,said capillary tube outlet means being thus operable to siphon out theentire content of liquid sample in said reservoir.

lill. Apparatus for analysis of multiple samples by thin layer and paperchromatography, which comprises A) a base for supporting achromatographic paper or plate; B) a plurality of spot applicator unitsmounted in side-byside relationship on said base, each spot applicatorunit comprising a. reservoir means for holding a measured amount of aliquid sample, said reservoir means having inlet means for filling saidreservoir means and capillary tube outlet means for siphoning saidmeasured amount of liquid sample out of `said reservoir means and onto apaper or plate on said base,isaid capillary tube outlet means having adischarge end outside the reservoir means and below the level of aliquid sample in said reservoir means;

b. air supply means for flowing a stream of low pressure air toevaporate solvent from said liquid sample siphoned out of said reservoirmeans and onto a paper or plate on said base; and

'c. support means for supporting said reservoir means and said airsupply means with said air supply means arranged to tlow `air over aliquid sample discharged from said discharge end, said support meansincluding adjusting means for adjusting the position of said air supplymeans relative to said discharge end of said capillary outlet meansthereby to control the rate of evaporation of solventfrom said liquidsample siphoned out of said reservoir means; and C) the support means ofsaidapplicator units being mounted in spaced realtionship on said base.l

12. Apparatus according to claim l1, wherein said air supply means hasaportion extending substantially vertically, said vertical portionhaving a flared outlet end positioned adjacent said discharge "end, theposition of said flared outlet end being adjustable by said adjustingmeans. l

13. Apparatus according to claim 12, wherein said reservoir means has anelongated body portion slightly inclined to the horizontal, and saidcapillary outlet means comprises a capillary tube having a rst part lthereof inside said elongated body pprtion and in communication with aliquid sample in said elongated body portion, and said discharge endlbeing provided by a v second part of the capillary tube positionedoutside of i posts are spaced apart from one another at regularintervals in a straight line.

18. Apparatus according to claim 11, wherein said adjusting meanscomprises a ball and socket universal joint.

19. Apparatus according to claim l 1, wherein said air supply meanscomprises an air tube for each unit, an air manifold, flexible air hosesconnecing said air tubes to said manifold, and air flow control means onsaid air hoses.

20. Apparatus according to claim 19, wherein said air flow control meansconsists of screw clamps.

2l. Apparatus according to Claim 19, wherein said air supply meanscomprise means to heat said manifold.

UNITED STATES PATENT OFFICE CERTIFICATE OF CORRECTION Patent No.3,766,884 Dated October 23, 1973 Invented@ Arthur Rosenthal ,p It iscertified that error appears in the above-identified patent and thatsaid Letters Patentare hereby corrected as shown below:

Cancel claim 10 and insert the following: i l0 The applicator unitaccording to claim 3,

wherein said inlet means for filling said reservoir is provided by avertical funnel-like tube projecting upwardly from said elongated bodyportion; and plug means are provided in one end of said elongated bodyportion, said capillary tube passing through .said plug means 24Apparatus according to claim 13, wherein said" inlet means for fillingsaid reservoir is provided by a vertical funnel-like tube projectingupwardly from said elongated body portion; and plug means are providedin one end of said elongated body portion, said capillary tube passingthrough sai plug means.

Claim 17, fline 2, "posts" should read support means Signed and sealedthis 17th day of September `1974.

(SEAL) "Attest:

t McCOY M. GIBSON JR. C. MARSHALL DANN y y Attestng Officer Commissionerof Patents FORM P01050 (10'69) UscoMM-Dc coen-ps9 U.S, GOVERNMENTPRINTJNG OFFCE: |959 0-36631,

1. A spot applicator unit for thin layer and paper chromatography, whichcomprises a. reservoir means for holding a measured amount of a liquidsample, said reservoir means having inlet means for filling saidreservoir means and capillary tube outlet means for siphoning saidmeasured amount of liquid sample out of said reservoir means, saidcapillary tube outlet means having a discharge end outside the reservoirmeans and below the level of a liquid sample in said reservoir means; b.air supply means for flowing a stream of low pressure air to evaporatesolvent from said liquid sample siphoned out of said reservoir means;and c. support means for supporting said reservoir means and said airsupply means with said air supply meAns arranged to flow air over aliquid sample discharged from said discharge end, said support meansincluding adusting means for adjusting the position of said air supplymeans relative to said discharge end of said capillary outlet meansthereby to control the rate of evaporation of solvent from said liquidsample siphoned out of said reservoir means.
 2. The applicator unitaccording to claim 1, wherein said air supply means has a flared outletend positioned adjacent said discharge end, the position of said flaredoutlet end being adjustable by said adjusting means.
 3. The applicatorunit according to claim 2, wherein said reservoir means has an elongatedbody portion, and said capillary outlet means comprises a capillary tubehaving a first part thereof inside said elongated body portion and incommunication with a liquid sample in said elongated body portion, andsaid discharge end being provided by a second part of the capillary tubepositioned outside of said body portion and arranged transverse to saidelongated body portion.
 4. The applicator unit according to claim 3,wherein said second part of said capillary tube is substantiallyperpendicular to said elongated body portion.
 5. The applicator unitaccording to claim 3, wherein said first part of said capillary tube isof stainless steel and said second part is of an insert plastic securedto said first part.
 6. The applicator unit according to claim 3, whereinsaid capillary tube is removably mounted in said elongated body portion.7. The applicator unit according to claim 1, wherein said air supplymeans comprises air flow control means.
 8. The applicator unit accordingto claim 1, wherein said air supply means comprises means for supplyingheated air.
 9. The applicator unit according to claim 1, wherein saidcapillary tube outlet means comprises a capillary tube having a firstpart inside the reservoir means and extending to the lowest part of saidreservoir means and a second part outside the reservoir means andextending below the level of said lowest part of said reservoir means,said capillary tube outlet means being thus operable to siphon out theentire content of liquid sample in said reservoir.
 10. The applicatorunit according to claim
 3. wherein said inlet means for filling saidreservoir is provided by a vertical funnel-like tube projecting upwardlyfrom said elongated body portion; and plug means are provided in one endof said elongated body portion, said capillary tube passing through saidplug means.
 11. Apparatus for analysis of multiple samples by thin layerand paper chromatography, which comprises A) a base for supporting achromatographic paper or plate; B) a plurality of spot applicator unitsmounted in side-by-side relationship on said base, each spot applicatorunit comprising a. reservoir means for holding a measured amount of aliquid sample, said reservoir means having inlet means for filling saidreservoir means and capillary tube outlet means for siphoning saidmeasured amount of liquid sample out of said reservoir means and onto apaper or plate on said base, said capillary tube outlet means having adischarge end outside the reservoir means and below the level of aliquid sample in said reservoir means; b. air supply means for flowing astream of low pressure air to evaporate solvent from said liquid samplesiphoned out of said reservoir means and onto a paper or plate on saidbase; and c. support means for supporting said reservoir means and saidair supply means with said air supply means arranged to flow air over aliquid sample discharged from said discharge end, said support meansincluding adjusting means for adjusting the position of said air supplymeans relative to said discharge end of said capillary outlet meansthereby to control the rate of evaporation of solvent from said liquidsample siphoned out of said reservoir means; and C) the support means ofsaid applicator units being mounted in spaced realtionshIp on said base.12. Apparatus according to claim 11, wherein said air supply means has aportion extending substantially vertically, said vertical portion havinga flared outlet end positioned adjacent said discharge end, the positionof said flared outlet end being adjustable by said adjusting means. 13.Apparatus according to claim 12, wherein said reservoir means has anelongated body portion slightly inclined to the horizontal, and saidcapillary outlet means comprises a capillary tube having a first partthereof inside said elongated body pprtion and in communication with aliquid sample in said elongated body portion, and said discharge endbeing provided by a second part of the capillary tube positioned outsideof said body portion and extending downwardly.
 14. Apparatus accordingto claim 13, wherein said second part of said capillary tube issubstantially perpendicular to said elongated body portion. 15.Apparatus according to claim 13, wherein said first part of saidcapillary tube is of stainless steel and said second part is of an inertplastic secured to said first part.
 16. Apparatus according to claim 13,wherein said capillary tube is removably mounted in said elongated bodyportion.
 17. Apparatus according to claim 11, wherein said posts arespaced apart from one another at regular intervals in a straight line.18. Apparatus according to claim 11, wherein said adjusting meanscomprises a ball and socket universal joint.
 19. Apparatus according toclaim 11, wherein said air supply means comprises an air tube for eachunit, an air manifold, flexible air hoses connecting said air tubes tosaid manifold, and air flow control means on said air hoses. 20.Apparatus according to claim 11, wherein said air flow control meansconsists of screw clamps.
 21. Apparatus according to claim 11, whereinsaid air supply means comprise means to heat said manifold. 22.Apparatus according to claim 11, including and means to heat said base.23. Apparatus according to claim 11, wherein said capillary tube outletmeans comprises a capillary tube having a first part inside thereservoir means and extending to the lowest part of said reservoir meansand a second part outside the reservoir means and extending below thelevel of said lowest part of said reservoir means, said capillary tubeoutlet means being thus operable to siphon out the entire content ofliquid sample in said reservoir.
 24. Apparatus according to claim 13,wherein said inlet means for filling said reservoir is provided by avertical funnel-like tube projecting upwardly from said elongated bodyportion; and plug means are provided in one end of said elongated bodyportion, said capillary tube passing through said plug means.